Combinatorial chemistry has become a powerful tool for drug discovery in the pharmaceutical and biotechnology industries. Generally speaking, combinatorial chemistry is defined as the repetitive and systematic covalent attachment of different structural moieties to one another to produce a mixture of numerous distinct molecular entities or target molecules (i.e., combinatorial libraries); desired target molecules include peptides, oligonucleotides, and small organic molecules. Frequently, combinatorial chemistry is utilized to generate a group of structurally related analogs which can then be evaluated to establish structure-activity relationships (SAR) and to optimize biological potency. See, e.g., M. A. Gallop et al., J. Med. Chem., 37:1233-1248 (1994)!.
Solid supports (e.g., polystyrene resin beads and silica chips) and, concomitantly, solid phase synthesis techniques are routinely utilized in generating combinatorial libraries. Unfortunately, presently used techniques are associated with several limitations that restrict their efficiency and widespread application. One major limitation is the difficulty of producing target molecules that, following cleavage from the solid support, provide diverse functional groups at the carboxamide of the target molecules.
Researchers have had some recent successes in the development of chemical techniques that allow the production of a target molecule with particular functional groups at the carboxamide terminus following cleavage from the support. One such success relates to the generation of target molecules bearing sulfonamide functional groups. Previously, the use of linkers possessing a sulfur atom anchoring the target molecule to the resin (e.g., the so-called "safety-catch" linker see, e.g., G. W. Kenner et al., J. Chem. Soc. Chem. Commun. 636 (1971)!) yielded only an acid or amide following cleavage of the sulfonyl peptide. C. Gennari et aL, Angew. Chem. Int. Ed. Engl. 34:1763 (1995)!. However, new techniques involving the coupling of sulfonyl chlorides to an amino functionalized resin afford target molecules possessing a sulfonamide group at the carboxamide terminus of the molecules after cleavage. K. A. Beaver et al., Tetrahedron Letters 37(8):1145-48 (1996)!.
Despite recent developments in the field of solid-phase organic synthesis applications see, e.g., Y. Han et al., J. Org. Chem. 61:6226-6339 (1996); T. L. Boehm et al., J. Org. Chem, 61:6498-6499 (1996)!, there is presently no efficient technique for producing target molecules possessing an N-alkyl amide (e.g., RCONHR') at the carboxamide termini of the molecules following cleavage from the solid support. Indeed, the only method routinely used for the solid-phase synthesis of N-alkyl amides involves the aminolysis of resin-bound esters. However, this procedure requires an excess of amine for the aminolysis step see, e.g., A. F. Abdel-Magid et al., J. Org. Chem. 61:3849-3862 (1996)!, necessitating a subsequent purification procedure. Moreover, when utilizing this procedure, the target molecule must be anchored to the solid phase by an ester linkage rather than an amide linkage; unfortunately, the acid- and base- sensitivity of the ester limits the types of chemistry which can be used to modify the tethered target molecule.
Rink's amide resin has been extensively used for the solid-phase synthesis of peptide fragments, as it affords carboxamide primary amides (RCONH.sub.2) upon cleavage from a substituted benzyhydryl amine linked to a polystyrene resin. H. Rink, Tetrahedron Letters 28:3787-3790 (1987)!. Although the Rink amide resin is useful for the synthesis of primary amides see, e.g., G. C. Look et al., Tetrahedron Letters 36:2937-2940 (1995)!, there are no reported methods for the direct alkylation of the Rink amide resin or amine resin to facilitate the incorporation of a secondary amide, thus yielding concomitant diversity at the resin-bound position.
Resin-bound amines distinct from Rink's amine resin have been alkylated by a reductive amination procedure. For example, Mokotoff et al. J. Med. Chem. 35:4696-4703 (1992)! reported the reaction of benzhydrylamine (BHA) resins directly with Boc-leucinal (in DMF containing 1% acetic acid), followed by reduction utilizing a solution of sodium cyanoborohydride (in DMF containing 1% acetic acid). However, the Mokotoff et al. procedure does not efficiently render N-alkylated amides in appreciable yields.
What is needed are methods and compositions for generating molecules containing a nitrogen-substituted amide at their amide termini (ie., that end of the molecules bound to the support) upon cleavage from a solid support. Such methods and compositions should allow for the production in high yields of target molecules with particular functional groups at their amide (e.g., carboxamide) termini.